2003:

2001:

Identifying the Effects of Environmental Toxicant Exposure on miRNA-regulated Adrenomedullin in the Development of Preeclampsia
Principal Investigator: Kathleen M. Caron, Professor and Chair of the Department of Cell Biology and Physiology, SoM
Award: $30,000

Abstract
Preeclampsia (PE) is a hypertensive pregnancy syndrome affecting 8% of women causing high blood pressure, breathlessness, and seizure until delivery. PE results from the improper invasion of the fetal trophoblast cells and malformation of the maternal spiral arteries resulting in nutrient deprivation and unstable vasculature. Adrenomedullin (AM) is a vasodilating protein, important for the development of the spiral arteries and PE prevention. Dosage of AM is critical in maintaining the maternal-fetal interface for a successful pregnancy. Indeed, AM is susceptible to estrogenic regulation and our collaborator recently identified placental accumulation of the metal, cadmium, correlates with increased PE risk and decreased AM levels. Through the use of miRNA mapping software, 133 miRNAs were determined to bind at the AM 3’ untranslated region (UTR) resulting in AM degradation. Recent data confirms cadmium upregulates specific placental miRNAs, three of which can bind to the AM 3’UTR. Thus, we hypothesize that exposure to environmental toxicants results in the expression of a unique group of miRNAs targeting AM for degradation, resulting in increased risk for PE. We will determine the effect of environmental toxin exposure on the expression of AM signaling components. Additionally, we will identify the unique population of AM-targeting miRNA species expressed upon treatment of human trophoblast cells with the phytoestrogen, coumestrol, or cadmium. These results can deepen our understanding of AM regulation to treat and prevent hypertensive pregnancy disorders.

AbstractAnimal models strongly suggest prenatal exposure to endocrine-disrupting chemicals (EDCs) may increase risk of obesity and related disorders by “re-programming” metabolism, but human data are limited and mixed. Recent studies point especially to the organochlorine compound DDE as a candidate obesogen. The proposed project will use a birth cohort in which fetal exposure to DDE was associated with rapid infant weight gain to address critical questions. We will conduct analyses to examine whether associations persist with overweight at age 5y, exploring complex issues such non-linear dose response, and whether associations may be attributable to chemical mixtures or postnatal exposure, rather than prenatal DDE exposure alone. We also aim to explore associations with early cardiometabolic markers, which have not been previously studied. This innovation will deepen understanding of whether EDCs may directly affect metabolism, vs. secondary to prolonged obesity. Results will inform critical design issues for an R01 application, by identifying: (i) cardiometabolic markers for study in a larger sample; (ii) candidate locus-specific epigenetic markers for study based on cardiometabolic markers affected; and (iii) approaches for studying effects involving exposure mixtures.

AbstractOrganophosphate pesticides, bisphenol A (BPA) and phthalates have been implicated in the etiology of Attention-Deficit Hyperactivity Disorder (ADHD), but there have been no prospective studies of prenatal exposure in relation to a clinically defined phenotype. We propose to investigate these relationships in the ADHD substudy of the Norwegian Mother and Child Birth Cohort. This pilot application seeks support for preliminary studies of the suitability of stored biospecimens within the MoBa biorepository to investigate our aims. We propose to conduct a reliability study of free and conjugated BPA over the course of pregnancy using stored MoBa urine samples; to examine the consequences of MoBa maternal plasma shipping in storage methods on FT4 concentration using a small population of women enrolled at UNC; and to investigate the feasibility of extracting FT4 from fetal blood spots. To achieve our aims we will be collaborating with the UNC-CEHS Biostatistics and Integrative Health Sciences and Facilities Cores. This application represents a transdisciplinary collaboration between behavioral scientists, epidemiologists, and clinical investigators, and addresses the complex interaction between environmental exposures and human neurodevelopment.

Epigenetics of the nutrient-carcinogen interactions: the role of choline in transplacental carcinogenesis associated with exposure to arsenic
Principal Investigator: Zuzana Drobna, PhD; Research Assistant Professor, Department of Nutrition
Award: $25,000

AbstractSeveral nutrients and certain environmental carcinogens have been shown to modify DNA methylation pattern during the fetal development. However, a little is known about the in utero interactions between these nutrients and carcinogens and how these interactions affect the methylation of genomic DNA or specific genes. Inorganic arsenic (iAs) is a potent human carcinogen. Chronic exposure to arsenic has been associated mainly with skin, liver, lung and bladder cancers. Animal studies show that in utero exposure to iAs may be in part responsible for these effects. Exposures of pregnant CD1 mice to iAs in drinking water from gestation day GD8 to GD18 have been shown to markedly increase liver tumor incidence and multiplicity in their adult offsprings. Results from this and other laboratories confirmed that prenatal exposures of mice to iAs alter methylation and expression of several genes associated with a variety of human and rodent tumors. In addition, we observed significant changes in the concentration of S-adenosylhomocysteine (SAM) in the livers isolated from fetuses exposed in utero to iAs. These findings indicate that exposure to iAs alters DNA methylation by competing with DNA methyltransferases (DNMT) for SAM, the donor of methyl groups used for methylation of both DNA and iAs. Alternatively, iAs and/or its metabolites may modify the expression or activity of DNMTs, thus, altering the DNA methylation pattern. Choline is a precursor for SAM synthesis. Choline intake has been shown to modify SAM availability. The proposed project will use the mouse model for transplacental arsenic carcinogenesis to examine effects of choline supplementation on epigenetic events that may determine the susceptibility of adult offsprings to cancer. Specifically, effects of choline intake on the concentration of SAM, metabolism of iAs, methylation of CpG-islands in genomic DNA and gene expression will be examined in the livers of CD1 mouse fetuses during in utero exposure to iAs. Results of this work will be used to design a long-term cancer study that will examine the association between choline intake and epigenetic changes in fetuses exposed in utero to iAs, and development of liver cancer in adult offspring.

AbstractEvidence is accumulating to implicate environmental toxicants and oxidative stress in a complimentary role in osteoarthritis (OA). Epidemiological studies show positive associations between OA severity and blood lead levels as well as selenium insufficiency. The problem with our understanding of the influence of the environment is that the precise link between oxidative stress and OA remains elusive. This compels us to examine the effect of oxidative stress on joint pathology using a murine model of surgically-induced knee instability, to determine if the time-course of disease is altered in animals maintained on a selenium-deficient diet. We will also examine the role of the transcription factor Nrf2, critically important for expression of antioxidant enzymes, by following OA in animals that lack Nrf2, and normal animals fed a potent activator of Nrf2. We hypothesize that oxidative stress will accelerate tissue damage, and conversely that enhancement of antioxidant gene expression will slow progression as observed by MRI and histolopathogy. The natural extension of this pilot data is to explore the mechanisms of heavy metal exposures as oxidant stressors in the progression of OA.

STAT3: An inhibitory factor on obesity-associated inflammation and insulin resistance

AbstractObesity-associated proinflammatory state and insulin resistance play an important role in the development of metabolic syndrome. STAT3 is a transcription factor that mediates the signaling of proinflammatory cytokines, induces cell cycle progression, and prevents apoptosis. This proposal tests the hypothesis that STAT3 in adipose tissue regulates the expression of proinflammatory genes and STAT3 deficiency in this tissue results in the inhibition of proinflammatory state, thus preventing insulin resistance. This hypothesis is based on our preliminary data showing that the ASKO mice are obese, but not diabetic when fed a low-fat diet. The specific aims are to: 1) determine the inflammatory status of low-fat diet fed ASKO mice; 2) determine whether a high-fat diet induces a proinflammatory state and insulin resistance in ASKO mice; and 3) determine what genes are affected in the adipose tissues of high-fat diet fed ASKO mice. This study will generate important pilot data for a future R01 proposal. It will help to understand the interaction between obesogenic environmental factors and developmental determinants of obesity and metabolic syndrome and to inform studies of new therapeutic approaches.

AbstractAccumulating toxicological data suggest that exposure to metals results in skeletal and joint toxicity. Yet, little attention has been directed to metals in relationship to osteoarthritis (OA), the most common cause of arthritis, characterized by profound bone and cartilage disruption. The primary objective is to understand how toxic metals (lead [Pb] and mercury [Hg]) affect OA. This proposal leverages bio-specimens and data from The Johnston County Osteoarthritis Project, a population-based prospective cohort of OA in African-American and White adults, in which associations between Pb, Hg, and selenium (Se) and OA were seen. Using linear regression and analysis of covariance, we will investigate associations between whole blood Pb and toenail Hg and 5 biomarkers of joint tissue metabolism related to OA pathophysiology in 376 men. Together with extant data on 375 women, we will examine factors likely to interfere with mechanisms of metal-associated OA (age, sex, and race) and identify factors (e.g., Se) that may counteract adverse responses to metal exposures. Further, this proposal will provide preliminary data for planned NIH proposals to examine additional metals in OA.

AbstractPrevious studies have shown that prenatal dieldrin exposure alters development of both the GABAergic and serotonergic (5-HT) systems [reviewed by (Lauder and Liu, 1998)]. Consequently, such prenatal exposure may enhance a diathesis for psychopathology characterized by deficient GABAergic and serotonergic neurotransmission (e.g., affective disorders like anxiety and aggression). This hypothesis will be tested using a mouse model for anxiety/aggression to determine the effects of prenatal dieldrin exposure on behavior and brain neurochemistry. Relative to ICR mice selectively bred for low aggression (NC100), high aggression mice (NC900) reproduce traits associated with human anxiety disorders. Importantly, the higher levels of anxiety and aggression observed in the NC900 mice are negatively correlated with markers of GABAergic and serotonergic function (Nehrenberg, 2004; Nehrenberg et al., 2005). Behaviorally, unselected ICR animals show intermediate (i.e., “normal”) levels of anxiety and aggression relative to NC900 and NC100 mice. This animal model represents a unique tool for determining whether individuals at risk for developing affective disorders may be more susceptible to the persistent effects of dieldrin exposure during prenatal development.

Genetic and early life origins of obesity, metabolic and cancer-related risk factors among FilipinosPrincipal Investigator: Linda Adair, Ph.D., Professor and Associate Chair, Department of Nutrition

AbstractThe proposed research focuses on the identification of early life exposures, genetic main effects, and gene-environment interactions contributing to the development of obesity and other components of the metabolic syndrome among Filipino young adults. We use detailed, prospectively collected data from The Cebu Longitudinal Health and Nutrition Survey, an ongoing sociodemographic study to which we will add blood collection for analysis of DNA and plasma biomarkers of CVD risk. The sample includes about 2000 individuals born in 1983-84, and their mothers. This rich data set allows us to examine interactions of prenatal and early life nutritional exposures, selected genotypes, and diet and physical activity as determinants of outcomes. We also include genetic and phenotypic data on the mothers of young adults. Our ultimate research goal is to model a wide range of obesity-related phenotypes reflected in weight gain trajectories from the prenatal period to adulthood, subcutaneous fat patterning, other markers of central obesity, and a suite of biomarkers of CVD risk (insulin resistance using HOMA, fasting glucose and insulin, leptin, C-reactive protein and other markers of inflammation, and serum lipids).

AbstractLong chain fatty acids contribute to the structure and function of brain and retina during normal development, which begins in gestation and continues through early childhood. Clinical trials have indicated that supplementing infant formula with specific fatty acids may improve visual acuity and cognitive function, especially among preterm infants. Little is known, however, about whether variation in the maternal fatty acid profile during pregnancy might affect the infant’s neurodevelopment. We propose to examine the relation between maternal fatty acid levels during pregnancy and the infant’s visual and cognitive development. This study takes advantage of the Pregnancy, Infection, and Nutrition (PIN) Study and the PIN Postpartum Study. PIN collects considerable information on the women during pregnancy, including diet, stress, infection, and blood samples. PIN Postpartum will conduct a home visit with the women 3 and 12 months after delivery to assess postpartum diet, physical activity and psychosocial factors. Children born to participants of the PIN Postpartum Study will be eligible to participate in the PIN Pediatric Study. This study will add an evaluation of the child’s development to the mother’s scheduled postpartum home visit. The developmental evaluation will include a test of visual acuity using Teller Acuity Cards, assessment of mental and motor skills using the Bayley Scales of Infant Development, and assessment of early language and communication skills by the MacArthur Communicative Development Indices. This study will also analyze the maternal prenatal fatty acid profile from erythrocyte samples that were collected during pregnancy and stored by the PIN Study. We will evaluate the variability in fatty acid levels and their correlation with reported maternal diet. We will conduct a preliminary investigation of the relation between maternal fatty acid levels and child’s neurodevelopment. This preliminary pilot data will be used to apply for NIH funding of a larger study that will distinguish more subtle associations between fatty acids and neurodevelopment. In the mean time, the outcome data generated will allow an efficient investigation of the association between neurodevelopment and other maternal exposures, such as smoking, infection, and stress, which already exist among PIN data.

AbstractThe aim of this project is to investigate gene-environment interactions in the etiology of developmental disabilities, like autism, using a mouse model for Smith-Lemli-Opitz Syndrome (SLOS). This project tests the general hypothesis that individuals with genetic predisposition to low cholesterol due to mutations in delta 7-dehydrocholesterol reductase (Dhcr7) may be at increased risk for abnormal brain development following prenatal exposure to neurotoxicants that target neurotransmitter systems. This hypothesis is based on: 1) evidence for vulnerability of developing GABA, serotonin (5-HT) and catecholamine neurons following exposure to organochlorine pesticides (e.g., dieldrin) that act as potent GABAA receptor antagonists; and 2) evidence for aberrant development of 5-HT neurons in mice with a targeted mutation in Dhcr7. The Specific Aims test the hypothesis that Dhcr7 mice will be more vulnerable to prenatal exposure to dieldrin or the GABAA receptor antagonist, bicuculline (positive control) than their wildtype counterparts. Pregnant mice will be exposed to different doses of pesticide or bicuculline from gestational day (GD) 11-16. In Specific Aim 1, dose-dependent treatment effects will be evaluated in adult offspring using a behavioral test battery to screen effective doses of prenatal treatments and determine long-term effects of prenatal exposure on social and perseverative behaviors. In Specific Aim 2, prenatal treatments will be limited to doses of pesticide or bicuculline that produce deficits in social behavior or promote perseverative responses. Affymetrix microarrays will be used to profile gene expression in brains of neonatal and adult offspring. Gene expression data will be validated using Real -Time PCR. Future studies will analyze proteins encoded by significantly altered genes using Western blotting or radiolabel immunocytochemistry on brain sections. Taken together, these studies should provide important insights into roles played by gene-environment interactions in the etiology of developmental disabilities, like autism, that involve deficits in social behavior and perseverative responses, and give clues to possible underlying cellular and molecular mechanisms.

AbstractThere is increasing evidence that infection and the inflammatory response may play a crucial role in the etiology of preterm birth (<37 wks). Intrauterine infection has been found to increase intra-amniotic and cervicovaginal fluid concentrations of inflammatory cytokines; and, both intrauterine infection and elevated inflammatory cytokine concentrations are associated with preterm birth. Consequently, it has been suggested that a woman’s own immune response to infection may precipitate preterm birth. Polymorphisms in the genes that encode the pro-inflammatory cytokines TNF-a and IL-1ß cause increased cytokine secretion in response to bacterial lipopolysaccharide (LPS) stimulation, resulting in higher inflammatory activity. Conversely, polymorphisms in cytokines IL-6 and IL-1Receptor antagonist increase anti-inflammatory activity. Many of these polymorphisms are relatively common in Caucasians (15%-47%), and have been associated with more severe outcomes in infectious and autoimmune diseases. In addition, the TNF-a (-307) polymorphism has been found to increase the risk of preterm birth in two relatively small studies. No other cytokine polymorphisms have been evaluated in relation to preterm birth.

We propose to examine the relationship between selected cytokine polymorphisms and risk of preterm birth in the large, well characterized, Pregnancy, Infection and Nutrition (PIN) cohort. DNA will be extracted from blood samples that have been obtained from 282 case and 618 non-case participants, and genotyping of three cytokine polymorphisms [TNF-a (-307), IL-1ß (-511), and IL-6 (-174)] will be performed. The main effect of individual polymorphisms and multiple polymorphisms on the risk of preterm birth will be examined. Finally, we will perform an exploratory analysis of the interaction between infection during pregnancy, cytokine polymorphisms, and preterm birth.

This project would provide a unique and efficient opportunity to examine genetic susceptibility factors for an important reproductive outcome. This preliminary research will permit a fuller exploration of a broader array of polymorphisms of other cytokines, other inflammation genes, and genes in other pathways. In addition, future work would include utilization of fetal DNA samples for genotyping. The project would engage members of the Developmental Susceptibility and Genetic Susceptibility Research Cores as well as utilize the High Throughput Genotyping and Biostatistics and Epidemiologic Methods Core Facilities.

AbstractUterine fibroids, or leiomyomata, are benign smooth muscle and connective tissue tumors. They are the most common pelvic neoplasm in women, affecting more than 20% of reproductive age women. Physiologic changes of pregnancy favor fibroid growth, however individual women may experience regression, stability, or growth of fibroids during gestation. Little is known about the natural history and mediators of growth of fibroids; even less is known about their etiology. We are interested in exploring the relationship between exposure to phthalates and presence and behavior of fibroids during pregnancy. Phthalates are ubiquitous man-made chemicals. Specific phthalates have hormonal activity in animal models and in vitro; including acting as endocrine disruptors. Routes of exposure include inhalation and dermal absorption from products such as perfumes, hairspray, soaps, skin care products, nail polish, and other cosmetics. It has recently become feasible to measure urine metabolites of common phthalates. This has allowed new research on the health effects of phthalates in humans and interest in methods to assess phthalate exposure in large-scale studies. Fibroids and phthalates are stated research priorities of multiple federal funding agencies. Our team proposes to lay a foundation for future work on phthalates and fibroids by: (1) developing an instrument to relate self-report of product use and other phthalate exposures to urinary phthalate levels; (2) obtaining self-report of use of products likely to contain phthalates from 100 women with, and 100 women without fibroids at baseline; (3) conducting follow-up ultrasounds at 22-24 weeks gestation for 40 women with fibroids to measure their fibroids; (4) measuring urinary phthalate levels for the first 30 women with fibroids who have follow-up ultrasounds and for 20 women without fibroids. This work will provide the requisite preliminary data and methodologic experience to guide continued research on fibroids, phthalates, and their potential influence on reproductive health outcomes.